Since early 1970s, the electroporation has been used to insert molecules into animal cells or plant cells. It is proved by researchers that when a cell is exposed to an instantaneous high-voltage pulse electric field, the cell membrane permeability increases due to a local fracture of cell membrane caused by the high voltage electric field such that pathways will be formed through the cell membrane, these pathways being referred as “electropore”. Although the existing times of these pathways are brief, it is enough to satisfy the requirement of the macromolecules such as proteins or plasmid DNAs entering or outletting. The cell may tolerate the high voltage which is used to format these pathways, however, the cell may be killed when these pathways are formed if the voltage of the high-voltage pulse is too high, the lasting time of the electric field is too long, or the times of the high-voltage pulse electric field is too many.
At the earlier, the electroporation is carried out by using two parallel-plate electrodes fixed on two inside walls of a container respectively. The cell suspension prepared for electroporation and the molecules which are expected to be introduced into the cell are mixed, and the solution is added into the electroporation container and placed between the two electrodes. In order to improve the effect of cell electroporation, an instantaneous high-voltage pulse is applied to the electrodes by one or more times so as to apply a high-voltage electric field pulse on the cell suspension between the electrodes. However, the distance between the parallel-plate electrodes is large, the required voltage is usually up to several hundred or even several thousand of volts, causing security and reliability issues, and the generation of a cathode effect is unavoidable, which has a huge damage to the cells. The planar electrode arose later reduces the distance between electrodes, and can generate an equal electric field intensity under a lower voltage and brings well electroporation effects, but it deals with a small amount of cells every time and is completely not suitable for high throughput experiment operations.
There is an electroporation instrument employing a three-dimensional electrodes on the market, but it usually uses for electroporations in clinic such as for tumour tissues or living tissues, etc. This type of instruments has a small number of electrodes and a simple combination, and some of them even use two needle-like electrodes as the three-dimensional electrodes, which meet the requirement of a living body and is easy to penetrate into the tissue and the living body, however, it's hard to be used for experiment operations of electroporation on extracorporeal cells such as suspended cells or attached cells.
At the current market, the most common electroporation container has a small volume, and needs multiple times of repeated operations during electroporation. Although this repetition of adding samples into the container for electroporation is easy and convenient, this container only can satisfy the requirement of a small scale cell electroporation for researchers and is not suitable for high throughput cell electroporations. In this method, it is impossible to keep sterile and it can not meet the requirement of a large volume of cell electroporation, and adding samples repeatedly will lengthen the actual operation time. Those issues are adverse to the accomplishment of the experiment.
In 1980s, researchers started to research the flow electroporation experiment methods used in treating the large volume of cells. Generally, the flow electroporation employs adapted parallel-plate electrodes and the cell suspension required to conduct electroporation flows continuously and steadily through between the two electrodes until the entire cell suspension is carried through electroporation such that the electroporation for a large volume of cells is achieved. When the cell suspension steadily flows through the two electrodes, the cells will be exposed to a high electric field pulse which is provided continuously at a constant interval. The flow electroporation method comprises an electroporation chamber with openings for the electrodes and cell suspension in and out. However, according to the hydrodynamics laws, when the fluid flows through the pathway between the parallel plates, the fluid in the middle of the pathway and the fluid at the periphery of the pathway have a difference in the flow speed, and the flow speed of the fluid in the middle of the pathway is greater than the flow speed of the fluid at the periphery of the pathway; the smaller the size of the pathway is, the faster the flow speed is and the more obvious this effect is. This kind of effect of fluid will introduce a shear force which may cause damages to the cells and goes against to the experimental process of electroporation. For the throughput of experiment, it is desired that the volume between the parallel plates is the larger the better, which may be achieved by enlarging the distance between the electrodes; for the applied voltage, it is desired that the distance between the parallel plates is the smaller the better, which may be achieved by reducing the pulse voltage and decreasing the cathode effect; for hydrodynamics lows, it is desired that the distance between the parallel plates is not so small, and the shear force is required to be reduced to a small level which is small enough to have no damage to the cells. Therefore, although using the parallel plates to design a flow electroporation chamber is relatively simple, there are many limits.